Gynecologic Oncology Center, Peking University People's Hospital, Beijing 100044, P.R. China.
Laboratory Animal Center, Peking University People's Hospital, Beijing 100044, P.R. China.
Mol Med Rep. 2014 Mar;9(3):955-60. doi: 10.3892/mmr.2013.1876. Epub 2013 Dec 20.
The aim of the present study was to produce transgenic mice expressing tumor virus A (TVA) in the ovary under ovarian specific promoter 1 (OSP1) control. A transgenic mouse model was established in which TVA, an avian retroviral receptor gene driven by OSP1, was selectively expressed in the ovary. A recombinant plasmid containing TVA cDNA and an OSP1 promoter was constructed. The DNA fragment was repeatedly injected into male mouse testes at multiple sites. At 4‑7, 7‑10 and 10‑13 weeks following the final injection, two DNA‑injected male mice were mated with four wild‑type female mice to produce transgenic mice. The transgenic positive rate in mouse F1 offspring was 39.69%. When the positive F1 individuals were mated with wild‑type Imprinting Control Region mice (PxW) or with positive F1 individuals (PxP), the F2 individuals had a transgenic rate of 12.44%. The transgenic rates in the F1 offspring, produced following mating at the three time intervals, were 55.71 (39/70), 30.77 (4/13) and 18.75% (9/48), respectively. The transgenic rates of the F2 offspring decreased with the age of the F1 offspring, from 26.67% when PxP were mated at 6‑8 weeks of age to 6.52% when PxW were mated at 5‑6 months of age. The results indicate a high efficiency of gene transfer to F1 offspring using testis‑mediated gene transfer (TMGT). The transgenic rate in the F2 offspring was lower than that in the F1 offspring. The results reveal that TMGT is suitable for creating transgenic animals among F1 offspring. Semi‑quantitative reverse transcription-polymerase chain reaction results showed that TVA was expressed in the mice ovaries. The results demonstrate the importance of using the replication‑competent avian sarcoma‑leukosis virus long terminal repeat with a splice acceptor‑TVA system in ovarian tumorigenesis research.
本研究旨在利用卵巢特异性启动子 1 (OSP1) 控制，在卵巢中表达肿瘤病毒 A (TVA)，构建表达 TVA 的转基因小鼠。构建了含有 TVA cDNA 和 OSP1 启动子的重组质粒。将 DNA 片段反复注射到雄性小鼠睾丸的多个部位。在最后一次注射后 4-7、7-10 和 10-13 周，将两只注射了 DNA 的雄性小鼠与四只野生型雌性小鼠交配，以产生转基因小鼠。F1 代后代的转基因阳性率为 39.69%。当阳性 F1 个体与野生型印迹控制区小鼠 (PxW) 或阳性 F1 个体 (PxP) 交配时，F2 个体的转基因率为 12.44%。在三个时间间隔内交配产生的 F1 后代中，转基因率分别为 55.71%（39/70）、30.77%（4/13）和 18.75%（9/48）。随着 F1 后代年龄的增长，F2 后代的转基因率逐渐降低，从 6-8 周龄 PxP 交配时的 26.67%降至 5-6 月龄 PxW 交配时的 6.52%。结果表明，利用睾丸介导的基因转移（TMGT）可高效地将基因转移到 F1 后代。F2 后代的转基因率低于 F1 后代。结果表明，TMGT 适合于在 F1 后代中创建转基因动物。半定量逆转录-聚合酶链反应结果显示 TVA 在小鼠卵巢中表达。结果表明，在卵巢肿瘤发生研究中使用具有剪接受体-TVA 系统的复制型禽肉瘤-白血病病毒长末端重复序列非常重要。
